Dale E C, Ow D W
Plant Gene Expression Center, U.S. Department of Agriculture/Agricultural Research Service, Albany, CA 94710.
Proc Natl Acad Sci U S A. 1991 Dec 1;88(23):10558-62. doi: 10.1073/pnas.88.23.10558.
A general method of gene transfer that does not leave behind a selectable marker in the host genome is described. A luciferase gene was introduced into the tobacco genome by using the hygromycin phosphotransferase gene (hpt) as a linked selectable marker. Flanked by recombination sites from the bacteriophage P1 Cre/lox recombination system, the hpt gene was subsequently excised from the plant genome by the Cre recombinase. The Cre-catalyzed excision event in the plant genome was precise and conservative--i.e., without loss or alteration of nucleotides in the recombinant site. After removal of the Cre-encoding locus by genetic segregation, plants were obtained that had incorporated only the desired transgene. Gene transfer without the incorporation of antibiotic-resistance markers in the host genome should ease public concerns over the field release of transgenic organisms expressing such traits. Moreover, it would obviate the need for different selectable markers in subsequent rounds of gene transfer into the same host.
本文描述了一种不会在宿主基因组中留下选择标记的基因转移通用方法。通过使用潮霉素磷酸转移酶基因（hpt）作为连锁选择标记，将荧光素酶基因导入烟草基因组。hpt基因两侧是来自噬菌体P1 Cre/lox重组系统的重组位点，随后通过Cre重组酶从植物基因组中切除。植物基因组中Cre催化的切除事件是精确且保守的，即在重组位点没有核苷酸的丢失或改变。通过遗传分离去除编码Cre的基因座后，获得了仅整合了所需转基因的植物。在宿主基因组中不整合抗生素抗性标记的基因转移应能减轻公众对表达此类性状的转基因生物田间释放的担忧。此外，这将消除在后续几轮基因转移到同一宿主中使用不同选择标记的必要性。
